Activation of transcription factor STATS is a defining feature of interleukin-6 (1L-6) family cytokines. Both IL-6 and STATS are protective for cardiomyocytes and necessary for ischemia preconditioning. However, mechanisms regulating IL-6 cytokine signaling in cardiomyocytes are largely unexplored. We propose that the protective character of IL-6 cytokine signaling in cardiomyocytes is determined by 3 mechanisms: redox- regulation of the kinase JAK1 that activates STATS, phosphorylation of STATS on S727, and induction of SOCS3 that terminates IL-6 cytokine signaling. Moreover, we hypothesize that while JAK1 activation and preconditioning will not occur without proper redox status, mismatched STATS serine-tyrosine phosphorylation and SOCS3 induction may ultimately prove detrimental. To investigate regulatory mechanisms of IL-6-family signaling in cardiomyocytes, we propose 3 aims.
Aim 1 is to establish the importance of JAK1 redox-sensitivity for ischemic preconditioning. We will assess the impact of glutathione depletion in vivo on IL-6-family cytokine signaling and preconditioning. Mutagenesis and biochemical experiments will be performed to define the mechanism for JAK1 redox-sensitivity. We will also assess if in vivo delivery of a redox-insensitive JAK1 mutant enhances or restores preconditioning protection.
Aim 2 is to determine the importance of S727 phosphorylation in preconditioning and cardiac remodeling. Our working hypothesis is that S727 phosphorylation is important for cooperative transcription, and thus which genes STATS activates is affected by which serine kinases are activated. We will address that hypothesis by assessing importance of S727 phosphorylation in preconditioning-induced expression of protective genes. We will also determine by microarray which STATS-induced genes do not require S727 phosphorylation, and establish if those genes encode proteins implicated in adverse cardiac remodeling.
Aim 3 is to establish the role of SOCS3 in ischemic preconditioning. Because SOCSS-deficient mice die in utero, we have generated heart-targeted SOCS3 knockout mice. A closed-chest mouse myocardial ischemia-reperfusion model will be used to assess the short- and long-term consequences of SOCS3 deletion on cardiac remodeling. We contend that understanding how IL-6 cytokine signaling is regulated has great significance for exploiting the therapeutic potential of these cytokines and the phenomenon of preconditioning.
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